CD8 T-cell activation in mice injected with a plasmid DNA vaccine encoding AMA-1 of the reemerging Korean Plasmodium vivax

Relatively little has been studied on the AMA-1 vaccine against Plasmodium vivax and on the plasmid DNA vaccine encoding P. vivax AMA-1 (PvAMA-1). In the present study, a plasmid DNA vaccine encoding AMA-1 of the reemerging Korean P. vivax has been constructed and a preliminary study was done on its cellular immunogenicity to recipient BALB/c mice. The PvAMA-1 gene was cloned and expressed in the plasmid vector UBpcAMA-1, and a protein band of approximately 56.8 kDa was obtained from the transfected COS7 cells. BALB/c mice were immunized intramuscularly or using a gene gun 4 times with the vaccine, and the proportions of splenic T-cell subsets were examined by fluorocytometry at week 2 after the last injection. The spleen cells from intramuscularly injected mice revealed no significant changes in the proportions of CD8(+) T-cells and CD4(+) T-cells. However, in mice immunized using a gene gun, significantly higher (P<0.05) proportions of CD8(+) cells were observed compared to UB vector-injected control mice. The results indicated that cellular immunogenicity of the plasmid DNA vaccine encoding AMA-1 of the reemerging Korean P. vivax was weak when it was injected intramuscularly; however, a promising effect was observed using the gene gun injection technique.     

Systematic study of the genus Compsilura Bouché in Southeast and East Asia with morphological and molecular data (Diptera,Tachinidae)

Compsilura concinnata (Meigen) is one of the most famous, most polyphagous and most widely distributed tachinid flies (Diptera, Tachinidae) in the world. This species is well known as a biocontrol agent of some injurious pests of cultural and wild plants and has been introduced from Europe to the United States to control mainly the gypsy moth. Recently we found three new species very closely resembling C. concinnata from Southeast and East Asia: C. lobata sp. nov. (Japan and Thailand), C. malayana sp. nov. (Malaysia) and C. pauciseta sp. nov. (Japan and Taiwan). Additionally, C. samoaensis Malloch is treated as a junior synonym of C. concinnata based on the examination of the type specimen. The genetic differences in the mitochondrial COI gene data are examined to assess the accuracy of species delimitation of Compsilura. The male postabdominal characters of these species are illustrated. The piercing female postabdomen of C. concinnata is illustrated and compared to those of other members belonging to the Blondelia group including Blondelia Robineau-Desvoidy, Celatoria Coquillett, Eucelatoria Townsend and Vibrissina Rondani.     

Exendin-4 improves steatohepatitis by increasing Sirt1 expression in high-fat diet-induced obese C57BL/6J mice

The effects of exendin-4 on Sirt1 expression as a mechanism of reducing fatty liver have not been previously reported. Therefore, we investigated whether the beneficial effects of exendin-4 treatment on fatty liver are mediated via Sirt1 in high-fat (HF) diet-induced obese C57BL/6J mice and related cell culture models. Exendin-4 treatment decreased body weight, serum free fatty acid (FA), and triglyceride levels in HF-induced obese C57BL/6J mice. Histological analysis showed that exendin-4 reversed HF-induced hepatic accumulation of lipids and inflammation. Exendin-4 treatment increased mRNA and protein expression of Sirt1 and its downstream factor, AMPK, in vivo and also induced genes associated with FA oxidation and glucose metabolism. In addition, a significant increase in the hepatic expression of Lkb1 and Nampt mRNA was observed in exendin-4-treated groups. We also observed increased expression of phospho-Foxo1 and GLUT2, which are involved in hepatic glucose metabolism. In HepG2 and Huh7 cells, mRNA and protein expressions of GLP-1R were increased by exendin-4 treatment in a dose-dependent manner. Exendin-4 enhanced protein expression of Sirt1 and phospho-AMPKα in HepG2 cells treated with 0.4 mM palmitic acid. We also found that Sirt1 was an upstream regulator of AMPK in hepatocytes. A novel finding of this study was the observation that expression of GLP-1R is proportional to exendin-4 concentration and exendin-4 could attenuate fatty liver through activation of Sirt1.     

Genomic Shifts,Phenotypic Clines,and Fitness Costs Associated With Cold Tolerance in the Asian Tiger Mosquito

Climatic variation is a key driver of genetic differentiation and phenotypic traits evolution, and local adaptation to temperature is expected in widespread species. We investigated phenotypic and genomic changes in the native range of the Asian tiger mosquito, Aedes albopictus. We first refine the phylogeographic structure based on genome-wide regions (1,901 double-digest restriction-site associated DNA single nucleotide polymophisms [ddRAD SNPs]) from 41 populations. We then explore the patterns of cold adaptation using phenotypic traits measured in common garden (wing size and cold tolerance) and genotype–temperature associations at targeted candidate regions (51,706 exon-capture SNPs) from nine populations. We confirm the existence of three evolutionary lineages including clades A (Malaysia, Thailand, Cambodia, and Laos), B (China and Okinawa), and C (South Korea and Japan). We identified temperature-associated differentiation in 15 out of 221 candidate regions but none in ddRAD regions, supporting the role of directional selection in detected genes. These include genes involved in lipid metabolism and a circadian clock gene. Most outlier SNPs are differently fixed between clades A and C, whereas clade B has an intermediate pattern. Females are larger at higher latitude yet produce no more eggs, which might favor the storage of energetic reserves in colder climate. Nondiapausing eggs from temperate populations survive better to cold exposure than those from tropical populations, suggesting they are protected from freezing damages but this cold tolerance has a fitness cost in terms of egg viability. Altogether, our results provide strong evidence for the thermal adaptation of A. albopictus across its wide temperature range.     

Transaminase-catalyzed asymmetric synthesis of <Emphasis Type="SmallCaps">l</Emphasis>-2-aminobutyric acid from achiral reactants

Asymmetric synthesis of an unnatural amino acid was demonstrated by ω-transaminase from Vibrio fluvialis JS17. l-2-Aminobutyric acid was synthesized from 2-oxobutyric acid and benzylamine with an enantiomeric excess higher than 99%. The reaction showed severe product inhibition by benzaldehyde, which was overcome by employing a biphasic reaction system to remove the inhibitory product from the aqueous phase. In a typical biphasic reaction (50 mM 2-oxobutyric acid, 70 mM benzylamine and 2.64 U/ml purified enzyme) using hexane as an extractant, conversion of 2-oxobutyric acid reached 96% in 5 h whereas only 39% conversion was obtained without the product extraction.     

Mating factor linkage and genome evolution in basidiomycetous pathogens of cereals

Sex in basidiomycete fungi is controlled by tetrapolar mating systems in which two unlinked gene complexes determine up to thousands of mating specificities, or by bipolar systems in which a single locus (MAT) specifies different sexes. The genus Ustilago contains bipolar (Ustilago hordei) and tetrapolar (Ustilago maydis) species and sexual development is associated with infection of cereal hosts. The U. hordei MAT-1 locus is unusually large (approximately 500 kb) and recombination is suppressed in this region. We mapped the genome of U. hordei and sequenced the MAT-1 region to allow a comparison with mating-type regions in U. maydis. Additionally the rDNA cluster in the U. hordei genome was identified and characterized. At MAT-1, we found 47 genes along with a striking accumulation of retrotransposons and repetitive DNA; the latter features were notably absent from the corresponding U. maydis regions. The tetrapolar mating system may be ancestral and differences in pathogenic life style and potential for inbreeding may have contributed to genome evolution.     

Measurement of changes in membrane surface morphology associated with exocytosis using scanning ion conductance microscopy

The extent that vesicles maintain a distinct identity and morphology after fusing with the plasma membrane is controversial. We used scanning ion conductance microscopy to image changes in the surface membrane of adrenal chromaffin cells after stimulation of exocytosis with a high K(+) solution. Within several minutes after stimulation, punctate depressions, 100-600 nm wide, were noted from 16% of the cells. The depressions were not randomly distributed, but appeared in clusters of two or more within a approximately 1 microm(2) area and disappeared after several minutes. Increases in membrane surface area, consistent with the fusion and collapse of one or more vesicles into the surface membrane, were observed in 64% of the cells after high K(+) stimulation. Surface area increases did not occur if the high K(+) solution did not contain Ca(2+). We conclude that scanning ion conductance microscopy can be used to follow the time course of surface membrane changes resulting from exocytosis and endocytosis.